• Our Behavioral Assays

    The Animal Behavioral Core offers PIs at Cincinnati Children’s and collaborators at other research institutions access to a wide range of behavioral and nonbehavioral assays. The procedures are available for mice or rats, with test apparatus scaled accordingly. Many of our measurement protocols incorporate video tracking and photocell technology, and we use SAS programs to analyze and present the captured data.

    The following tabs describe the assays we offer. For more information, or to arrange a collaboration with the Animal Behavioral Core, contact us at 513-636-8622.

  • Available Assays

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    The Acoustic (ASR) and Tactile Startle Response (TSR) are well-established tests for evaluating the complex brainstem-mediated reflex pathway. We use the San Diego Instruments SR Apparatus for these tests.

    We also test for pre-pulse inhibition (PPI), a well-established measure for testing sensorimotor gating. PPI is abnormal in several human disorders, such as schizophrenia and major depression. ASR and PPI test methods and responses are nearly identical in humans and rodents, offering homologous comparisons across species.

    Active/two-way/shuttle box avoidance test using the San Diego Instruments two-sided Gemini System to test the ability of animals to learn to respond to a warning signal (light, tone, or both) that the grid floor is about to deliver a mild aversive stimulus (foot shock).  If the animal crosses to the opposite side from where they start, they avoid the shock; if they wait until the warning period is over, the shock is activated and they must escape to the opposite side.  Two-way active avoidance is a classically conditioned response where the signal is the conditioned stimulus (CS), the shock is the unconditioned stimulus (US), escape is similar to an unconditioned response (UR), and avoidance is a conditioned response (CR).  A typical test requires 20-25 trials/day for 5 days to obtain adequate learning.

    This test can be modified to fit a PI’s specific needs. Typically, the test involves familiarizing the test animal with a test chamber, and then observing changes to the animal’s behavior after it is injected with one of a variety of drug challenges. We typically test animals using indirect dopaminergic agonists, specific dopamine D1 and D2 agonists and antagonists, cholinergic-muscarinic antagonists and glutamatergic-NMDA receptor antagonists. We also have experience using a variety of other agents for this test.

    This uses a labyrinthine maze with nine T-shaped cul-de-sacs branching from a central (but circuitous) channel leading from the start to the goal. The test is conducted in complete darkness (monitored with an infrared light emitter and camera) to eliminate distal cues, thus testing the animal’s “sense of direction” or ability to navigate using route-based egocentric cues (i.e., internal cues).

    We record latency time to find the escape and errors of commission (entry into blind alleys).  The test provides a method for evaluating an animal’s entorhinal cortex, where grid cells integrate information from the hippocampus and the pre- and post-subiculum (where head direction cells are located). 

    This test uses cued conditioning to evaluate amygdala function and contextual conditioning to evaluate function of the hippocampus, subiculum, cingulate, prefrontal and perirhinal cortices. After being conditioned by pairing a neutral stimulus (a tone) to an aversive stimulus in a specific test environment it will associate the tone with the aversive stimulus.  The animal is placed in the test chamber a second time and observed for reactions to the chamber alone or an altered test chamber accompanied by the tones.

    This is a treadmill based walking analysis system from Mouse Specifics that uses a transparent belt on the treadmill and a high speed digital camera mounted beneath the treadmill to record the animal’s foot placement and stride characteristics.  The software provides a large number of measurements of different aspects of gait and can provide very detailed data.  The test is not suitable for screening but rather for in-depth gait analysis as training animals to walk only forward on the treadmill takes some time and rodents need to be tested at different walking and running speeds and in some cases when the treadmill is inclined.  The data extraction and analysis of the many variables provided takes time to sort and analyze.

    These tests use elevated plus (EPM) or elevated zero (EZP) mazes to test anxiety-like behavior. The mazes are similar, with two open and two  closed  arms or quadrants (closed sections have high, black acrylic walls). Animals are tested for five minutes, and the time spent in the open areas provides the principal index of anxiety. These tests are video tracked and computer scored using AnyMaze software from Stoelting Instruments.

    The Forced Swim Test (FST) is a well-known test for assessing stress responses including HPA axis markers such as ACTH, corticosterone and others. It also serves as a test of depression (the Persolt test of swimming despair). The Persolt test evaluates swimming immobility after a training session in which the animal is placed in a cylinder half-filled with water. We can score the FST using video tracking software.

    Passive avoidance uses the two-sided Gemini apparatus from San Diego Instruments.  For this test, one side is lighted and the other side dark with a door in between.  The animal is placed in the lighted side for a period of time, then the door is opened, and the animal is timed for how long it takes them to cross to the dark side.  Once they cross to the dark side, the door is closed and they are given a moderate, short, fixed-duration foot shock.  They are then removed and tested for retention at different intervals after the first trial to see how long it takes them the second time to reenter the dark compartment.  In some procedures trials are repeated until they learn to remain on the lighted side for 3 min whereas in 1-trial passive avoidance they are given only one training trail and retention is tested at some predetermined interval later to see how well they made the dark-shock association.  This test will show deficits in animals with hippocampal lesions if the lesion is of sufficient severity. 

    Using the equipment and conditioning designed for the Conditioned Fear test, we are able to evaluate the effect of pre-exposure to a stimulus on conditioning. Animal models of schizophrenia and autism show an impairment of latent inhibition, which mirrors human characteristics of those diseases.

    This test evaluates the time it takes the test animal to move from light to dark areas in a divided test chamber, and dwell time between the two chambers. The test uses our photocell-equipped locomotor activity test chambers.

    This fully automated test is conducted for various time intervals, but typically lasts 60 minutes. The system records horizontal activity, total distance traveled (cm), central versus peripheral movement or time, vertical movement, repetitive movements and time the test animal spends exploring different zones of the environment. The test measures exploration in response to a novel environment, habituation rate (as the environment become familiar) and baseline spontaneous ambulation. The locomotor activity test can provide data on the test animal’s basal ganglia (caudate-putamen) function.

    This is the most widely used test for measuring spatial navigation and reference memory. The test animal is placed in an open, circular pool in which there is a submerged platform. A series of tests can measure a variety of memory and navigation tasks, as the test animal swims to the platform from various locations, or tries to find the platform after it has been removed. Tests are video-tracked using AnyMaze, and analysis can provide information on the test animal’s hippocampus-dependent reference memory and prefrontal cortex working memory.

    This test uses the animal’s reaction to a novel object as an avenue to evaluate dorsal hippocampal function. The animal is familiarized with two identical objects, and then is placed in a test space with one of the familiar objects paired with a novel object. We measure time spent exploring the novel object, which reflects intact memory for the original object.

    This test places a grid of foreign objects (marbles) in a standard cage filled with bedding (measured to a standard depth). Animals are tested for 30 minutes, and the number of objects buried is recorded. This is a test of defensive anxiety.

    The Core has added a new test of rodent anxiety reported to be an improvement over other methods. The test involves placing animals on an elevated open platform with slanted panels on two sides. The animal is tempted to climb down these slanted surfaces to reach the floor and escape. The slanted panels are designed so the animal can grip to some extent but not easily, creating fear that it may fall. But the surfaces have enough traction that the animal can hold on if it ventures over the edge onto the sloping panels. Consequently, the animal spends most of its time at the edge trying to climb down. This maintains the animal in an extended state of anxiety. The degree of anxiety is inferred by measuring how much time it spends at the edge versus retreating from the edge to the center of the platform. A second advantage of this test is that animals can be assessed repeatedly unlike other tests of anxiety that rely on fear of a novel environment. The problem with novelty tests is that once the novelty wears off, the test no longer measures anxiety. The new method avoids this since the impulse to climb down the slanted surfaces is based on the urge to escape not on novelty.

    The RWM is a swimming version of the radial-arm maze (RAM) that uses food reward.  The Core has an 8-arm RWM.  The RWM, like the RAM, tests either spatial trial-dependent, working memory or a combination of spatial working and trial-independent reference memory.  In the classic RAM, rats must be food restricted and trained to eat some preferred food reward, then trained to find these rewards or baits within the maze until they learn to search all the arms and go to the very end of an arm to find the bait.  After training, testing begins.  For working memory, all arms are baited for each new trial.  The animal must find all 8 rewards without repeating an arm visit from which it already took one of the rewards.  The trial continues until all 8 rewards are retrieved or up to a maximum of 10 min.  In the mixed working/reference memory version some arms are baited anew each day and some are never baited.  In this version the animal’s task it to find the new rewards each day without repeating an arm entry and never enter the unbaited arms.  Our version is a modified version of this that avoids food deprivation and training.  One arm in the RWM is always the start and at the end of the other 7 arms is a hidden, submerged platform.  On the first trial the animal can enter any arm and reach a platform upon which it is removed and that platform is removed so that on the next trial there are only 6 remaining platforms.  The animal’s task is to find any other platform without reentering the one it found first.  This continues for 7 trials until each one is found and each reentry to an arm without a platform is counted as an error.  The task can be modified to be a mixed working/reference spatial memory task just as the RAM is.  The working memory version typically requires 10-14 days for animals to become proficient with the task.

    The apparatus is the San Diego Instruments Rotor-rod System.  The rotor-rod is a rotating drum whose speed is computer controlled.  Animals are first taught to walk on the rotating surface at a slow speed.  They are then tested in a series of trials in which the speed is increased at regular short intervals until they cannot keep up and fall.  How long and at what speed they can remain on the drum is a measure of motor coordination and to a lesser extent balance.

    This test places the animal in a three-compartment test chamber.  The two end compartments each contain a small cage with wide slits to allow an animal on either side of the cage to contact and smell a conspecific on the other side.  For the test, a “stranger” mouse or rat is placed in the cage inside one of the end compartments.  The test subject is placed in the middle compartment and allowed to explore freely.  The time the test subject spends with the cage with the conspecific in it versus time in the opposite end compartment with the empty cage reflects social preference for another animal of the same species and strain.  There is evidence that social preference in rodents has some relationship to human sociability.

    The original Star Maze has 5 arms.  Its purpose is to test which learning strategy and type of memory an animal uses to find a hidden platform.  Animals are first trained to swim from one specific arm to another where a hidden platform is located.  Animals are trained over repeated trials until their performance is nearly flawless.  Then they are given a test or probe trial in which they are started from a different arm.  If they are spatially navigating, then the start location will not matter and they will swim to the arm that is closest to where the distal cues are that they used to find the platform during training.  If they are using internal path cues (egocentric navigation) they will use the same series of right and left turns they used during training and end up in a different (but predictable) location.  If they use cues on the walls of different designs, they will end up in yet a different arm.  We have adapted the 5-arm design to an 8-arm but the concepts are the same.

    The test measures immobility in the animal by suspending it by the tail using an acrylic plate with a hole for the tail to pass through. Once the animal determines that escape is impossible, it stops struggling. This test can show the effect of antidepressants and sedatives on the animal, and is often used to provide converging evidence for results from Forced Swim Testing (FST). Animals are suspended for single trials of no more than six minutes, and the time spent immobile is manually recorded.  This test is available only for use with mice.

  • Contact Us

    For more information on collaborating with the Animal Behavioral Core, call us at 513-636-8622.